Cellular assays are critical tools in the drug discovery process and in basic research. In the future, these assays will play a major role in systems biology, permitting the examination of cell structure and function and the determination of a drug compound's ability to enter a cell, the compound's toxicity and its overall efficacy. Advances in imaging technologies, florescent probes, and assay automation are predicted to drive the worldwide cellular assays market from an estimated $300 million in 2002 to $500 million in 2007. The most common application for cellular assay technology in drug discovery is target validation and lead identification and optimization. However, the complexity and richness in cellular assay data sets, compared to genomics and proteomics studies, will provide scientists with unparalleled tools to aid discovery efforts throughout the drug discovery process as well as for basic research applications.
To achieve the goal of measuring the spectrum of molecular events in cells, there is definite need for “in cell sensor probes” that quantitatively measure protein (or other) activities directly in cells in a regulated fashion to give real time functional data, without using expression vectors. These “in cell sensor probes” will define pathways in a Parallel Quantitative Biology (PQB) format for systems biology and enable novel regulated cell-based functional screening in a high throughput mode. This invention provides these and other probes, termed PAC probes (PhotoActivated Cell probes) herein.
Just as RNA abundance does not always correlate to protein synthesis, protein abundance does not always correlate to a protein's activity and function within the cell. For example, depending on the protein's subcellular location (e.g. cytoplasm, nucleus, etc.), if it is membrane bound, or if it is bound to another molecule (e.g. aptamer, peptide, protein, RNA, DNA), its activity may or may not correlate to its abundance within the cell. In addition, how a drug binds to a protein in vitro and in vivo can differ significantly, not to mention that the drug may not enter the cell and, thus, may not be functionally active. The PAC probes of the invention measure activities in a regulated real-time fashion in the cell while co-locating, interrogating the function of proteins, pathways and networks and detecting the mode of action of drugs within the cells in a high throughput fashion. Thus, PAC probes enable functional proteomics and genomics.